This proposal seeks to make therapeutic advances by characterizing targeted therapy resistance mechanisms in gastrointestinal stromal tumors (GISTs). Most GISTs express mutant, constitutively activated, KIT or PDGFRA oncoproteins. We have shown that these formerly untreatable cancers can be palliated in 80% of patients by oral single-agent therapy with the KIT/PDGFRA inhibitor, imatinib. Patients who develop resistance to imatinib can benefit from second-line therapy with sunitinib, which is an alternate KIT/PDGFRA inhibitor. Ultimately, however, most GIST patients will progress on both of these FDA- approved kinase inhibitors. Therefore, the aims of the research proposed here are to characterize imatinib/sunitinib resistance mechanisms in GISTs, and then identify therapeutic strategies that can circumvent the resistance mechanisms. Notably, our preliminary studies show that GIST imatinib resistance mechanisms vary from patient to patient, and also between metastatic lesions in a given patient. We have shown that even a single progressing GIST metastasis contains subsets of cells whose imatinib resistance mechanism differs from those in other cells from the same tumor focus. In the present effort, by revealing the scope - and particularly the heterogeneity - of the imatinib/sunitinib resistance problem in GIST, we will provide the understanding needed to design more effective clinical strategies. At the same time, these studies will enable the development of biomarkers,assays and cell lines to enable preclinical validation of novel therapeutic strategies to circumvent imatinib/sunitinib resistance. The goal in these studies is to translate the understanding of imatinib/sunitinib resistance into improved medical therapy for GIST patients who are progressing on imatinib or sunitinib. Initially, we will evaluate HSP90 inhibition as a strategy to inhibit the varied gain-of-function KIT mutations that manifest, in each patient, at the point of clinical progression on imatinib and sunitinib. This will be accomplished through a phase l/ll clinical trial of the HSP90 inhibitor, IPI-504, combined with imatinib, in patients showing progression of metastatic GIST on imatinib or sunitinib. Through these studies, we will translate the basic science proposed in this SPORE through to clinical application.